In recent years, a variety of assay methods based on immunological reaction have been employed for assaying microlevel substances in samples. Such a variety of immunoassay methods include the RIA method, the EIA method, immunoturbidimetry, the latex agglutination method, the metal colloid agglutination method, and immunochromatography. Among them, the latex agglutination method, the metal colloid agglutination method, and the like are generally employed, since these methods require no separation or washing of reaction mixtures, which is advantageous for automated assay. In immunoassay, it is general that a suitable method according to the target substance concentration of the biological sample is selected. The latex agglutination method and the metal colloid agglutination enable more accurate determination of a substance in a microamount as compared with immunoturbidimetry, but the substance in a microamount cannot attain a microlevel assay that can be determined by the RIA method and the EIA method. In any of these immunoassay methods, there is demand for reduction in the scale of reaction system and shortening of measurement time, and enhancement in measurement sensitivity is an important issue.
One known technique for enhancing measurement sensitivity is addition of polyethylene glycol or a water-soluble polysaccharide such as dextran to the reaction system (Patent Documents 1 and 2). However, since these additives have a molecular weight distribution and are not a single compound, stable sensitivity-enhancing effect cannot be attained, which is one problem.
In any immunoassay method, non-specific reaction with various miscellaneous substances contained in a sample other than the target antigen-antibody reaction causes aggregation or absorption, which problematically results in a decrease in measurement precision. Particularly when a plasma-derived biological sample containing hemoglobin originating from hematocytes is assayed, hemoglobin may influence obtaining accurate measurements of immunoassay. Since the extent of hemolysis varies depending on sample, accurate measurements cannot be obtained. Thus, the influence of hemoglobin must be avoided by a certain technique.
Hitherto, there has been proposed a method for avoiding the influence of hemoglobin through addition of a surfactant to the reaction system (Patent Document 3). However, addition of a surfactant decreases measurement sensitivity in immunoassay, thereby failing to determine a target substance at high precision, which is problematic.
Patent Document 4 discloses an immunoassay method for CRP in a human serum sample by use of anti-human CRP rabbit serum, in which non-specific reactions which would otherwise be caused by a complement component can be prevented by adding a polyhydric phenol to the assay system. In one disclosed specific procedure, non-specific reaction which would otherwise be caused by a complement or the like can be successfully prevented by addition of sulfuric acid calix(6)arene in an amount of 5 mM. However, Patent Document 4 merely discloses the action of calixarene on the influence of complements in CRP assay of a human serum sample by use of anti-human CRP rabbit serum, and does not disclose how calixarene gives an effect on the measurement sensitivity or on the influence of hemoglobin. In addition, Patent Document 4 is silent on a method for assaying target antigen or antibody by use of an insoluble carrier (e.g., latex particles) sensitized with an antibody to the target antigen or an antigen to the target antibody.